5-Methylcytosine is a methylated form of the DNA base cytosine that is believed to be involved in transcriptional regulation. When cytosine is methylated, the DNA maintains the same sequence, but the expression of methylated genes can be altered.
The function of this chemical varies significantly among species: in bacteria, 5-methylcytosine can be found at a variety of sites, and is often used as a marker to protect DNA from being cut by native methylation-sensitive restriction enzymes; in plants, 5-methylcytosine occurs at CpG, CpHpG and CpHpH sequences (where H=A, C or T); and, in fungi and animals, 5-methylcytosine predominantly occurs at CpG dinucleotides. Most eukaryotes methylate only a small percentage of these sites, but 70-80% of CpG cytosines are methylated in vertebrates.
Cytosine methylation in vertebrates typically occurs at CpG sites (cytosine-phosphate-guanine sites, that is, where a cytosine is directly followed by a guanine in the DNA sequence). The formation of Me-CpG is catalyzed by the enzyme DNA methyltransferase. About 80-90% of CpG sites are methylated in human DNA, but there are certain areas, known as CpG islands, wherein none of the CpG dinucleotides are methylated. These are associated with the promoters of 56% of mammalian genes, including all ubiquitously expressed genes. One to two percent of the human genome are CpG clusters, and there is an inverse relationship between CpG methylation and transcriptional activity.
The method described herein provides a way to estimate the amount of a methylated locus in a sample.